The ability of proteins to self-assemble into macromolecules is fundamental to all living organisms. Some of the most complex of these structures are the fibrils formed from the triple helical collagen. The precise manner in which the monomers self-assemble into fibrils has not been defined. The present application is based on our observations that the telopeptides of collagen I bind to a specific region spanning amino acids 776 to 822 of the alpha1(1) chain, and that this interaction is critical for collagen self-assembly. Our recent results of the assembly of collagen I and collagen II mutants, strongly support our hypothesis that site-specific interactions between collagen molecules are critical for the fibril formation. We now use the gene-engineered collagen II and collagen IX to study the mechanism of the formation of homotypic and heterotypic fibrils in detail. The broad, long-term goal of this project is to understand fundamental principles of the formation of collagen fibrils in health and disease. The overall hypothesis is that the specific interaction between collagen monomers during fibril assembly is controlled by the binding of telopeptides to specific regions of the triple- helical domain. This binding is critical for the regulation of collagen fibril formation. Moreover, we hypothesize that assembly of heterotypic fibrils is also controlled by the site-specific formation. Moreover we hypothesize that assembly of heterotypic fibrils is also controlled by the site-specific interactions between different collagen types, and that mutations in fibrillar collagens change the site-specific interaction and, as a result, alter the structure of the extracellular matrix. To test the hypotheses, the following specific aims are proposed: (1) To elucidate a site-specific mechanism of the formation of homotypic and heterotypic collagen fibrils, (2) To determine how the site-specific interactions between collagen molecules regulate morphology of fibrils, kinetics and thermodynamics of collagen self-assembly, and (3) To analyze how mutations in fibrillar collagens affect the site-specific mechanism of collagen assembly. The proposed research will yield new information on collagen self-assembly structure of collagen fibrils. Results of the proposed studies will have a wide application in the areas where collagen fibrils play an important role. Examples include cell differentiations and proliferations, development, bone mineralization, tissue remodeling, fibrosis, tissue engineering, and therapy of the heritable diseases of connective tissue.